The concentration of cortisol, progesterone, testosterone and delta-androstenedione in porcine follicular fluid, peripheral blood, perifollicular blood and ovarian venous blood are being measured and the determination of estrogens in the above samples will be continued. The study of the binding of progesterone, testosterone and estradiol-17 beta to the albumin-free follicular fluid fraction from pig ovaries will be investigated. The cortisol binding protein in the follicular fluid has been characterized and isolated in pure form. Similarly, the other steroid binding protein(s) will be isolated by the affinity chromatography and its physicochemical properties will be studied. The role of such binding protein component in the process of ovulation will be investigated. This could be achieved by its direct administration into follicle or a specific antibody will be produced in rabbit and then used to block the action of steroid binding component. If such component(s) is found physiologically active, it will be quantitated by radioimmunoassay. The site of production of steroid binding protein in the follicular fluid will be investigated. First, the properties of such steroid binding protein from follicular fluid will be compared to that of blood plasma. If found similar, the binding protein will be labeled and injected into pig or rat and its appearance in the follicle will be studied. Secondly, granulosa cells will be cultured and the possible synthesis and release of steroid binding component or other protein fraction that is contributed in the follicular fluid will be investigated. Specific steroid cytosol and nuclear receptor in the granulosa cells will be examined to correlate the fraction of steroids on the development of follicles and ovulation.